Bone marrow transplant represents hope for survival for many patients who suffer from blood, lymphatic and bone-related disorders, and from depleted marrow cells resulting from intensive chemo- and/or radiotherapy. In an "allogeneic" bone marrow transplantation ("allo-BMT"), bone marrow is transplanted from a donor other than the patient (autologous transplant) or the patient's identical sibling (syngeneic transplant).
Allogeneic BMT is the treatment of choice for many hematologic malignancies, such as leukemia, lymphoma and multiple myolema (1, 2). It is the only curative therapy for chronic myeloid leukemia. Transplantation of allogeneic bone marrow, particularly when employed together with high-dose chemoradiotherapy, has been shown to produce superior results compared to autologous or syngeneic transplants (3).
The advantages associated with allogeneic BMT are limited by the risk of a potentially life-threatening complication, graft versus host disease (GvHD). In performing allogeneic BMT, the patient initially undergoes an immunosuppressive regimen to minimize rejection of the graft. Severe GvHD following allo-BMT can be controlled by: (i) treating the patient to eliminate residual recipient alloreactive T cells, and (ii) treating the graft to remove mature alloreactive T lymphocytes (2).
Removal of alloreactive mature T lymphocytes from the allo-BMT increases the incidence of disease relapse, graft rejection and reactivation of viral infection (4). To counteract these effects, allo-BMT patients have been treated by introducing donor T lymphocytes after a delay following allo-BMT.
Recent studies have shown that delayed introduction of donor T lymphocytes following allo-BMT is a promising therapy for reconstituting immunity and treating relapse of several disease states, and a therapy which could render allo-BMT more efficacious. Patients that underwent treatment with allo-BMT devoid of T lymphocytes, when affected by recurrence of chronic myelogenous leukemia, acute leukemia, lymphoma, and multiple myeloma, could achieve complete remission after the infusion of donor leukocytes, without requiring cytoreductive chemotherapy or radiotherapy (5). In other studies, delayed lymphocyte introduction was used to treat complications related to the severe immunosuppression associated with allo-BMT, such as Epstein-Barr virus-induced B lymphoproliferative disorders (EBV-BLPD) (6, 7) and reactivation of cytomegalovirus (CMV) infection (8).
The therapeutic promise of delayed introduction of donor T lymphocytes following allo-BMT, however, remains limited by GvHD, a frequent and potentially lethal complication of the treatment. Currently, no specific treatment exists for established GvHD. Thus, the threat of GvHD must be weighed heavily against the therapeutic effect of allo-BMT, and limits the applications in which the therapy is employed. (9). Accordingly, a regimen for preventing and for treating GvHD is highly desired in order to permit the beneficial use of delayed introduction of donor T lymphocytes following allo-BMT.
In this delayed lymphocyte introduction therapy, there also is a need for a simple method of monitoring the lymphocytes post-infusion. Effective monitoring would permit an investigator to determine whether the infused lymphocytes contribute to or cause a variety of complications which may occur after infusion. Since complications post-BMT can arise from a variety of origins and since the patients are highly immunosurpressed, rapid determination of the mechanisms underlying complications is highly desired.
Recently, investigators have transduced lymphocytes for delayed introduction with a single selectable marker, the gene, neomycin phosphotransferase (neo). Thereafter, PCR was employed to monitor the gene in cells biopsied from the patient. However, this method is cumbersome and PCR is time consuming (19). For example, prior to infusing the transduced markers into a patient, expression of neo must be verified. Expression of neo in transduced lymphocytes takes about two weeks time. During such time, the lymphocytes grown in culture often undergo self-tranformation and mutation of their original characteristics. Also, during such time, the recipient patient's clinical condition often changes for the worse.
Any vector carrinying a marker gene, to be useful in the present context is safe, efficient, and preferably must not substantially interfere with the lymphocyte's range of immune functions or its longevity (persistence) in the recipient's immune/circulatory system.
Hence, it is desired to provide a vector carrying a marker which permits efficient and fast expression, easy detection, by methods such as fluorescence activated cell sorting, easy monitoring after infusion (particularly of peripheral lymphocytes) and of course, is safe and nonimmunogenic. This would permit better, speedier, specific diagnosis of complications, and concommitantly, more successful treatment.